Multiparameter flow cytometric analysis of CD326 expression on BALB/c Mouse thymocytes and splenic T lymphocytes. Left Plot: Mouse thymocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) [Cat. No. 553141/553142] and stained with BD OptiBuild™ BV480 Rat Anti-Mouse CD4 (Cat. No. 746475) and BD Horizon™ BUV737 Rat Anti-Mouse CD8a (Cat. No. 612759) antibodies and with either BD Horizon™ RB780 Rat IgG2a, κ Isotype Control (Cat. No. 568698; dashed line histogram) or BD Horizon™ RB780 Rat Anti-Mouse CD326 antibody (Cat. No. 568736/568737; solid line histogram) at 1.0 μg/test. The histogram showing CD326 expression (or Ig Isotype control staining) was derived from CD4- and CD8-negative gated events with the forward and side light-scatter characteristics of viable thymocytes. Middle and Right Plots: Mouse splenic leucocytes were preincubated with Mouse BD Fc Block™ and stained with BD Horizon™ BUV395 Hamster Anti-Mouse CD3e (Cat. No. 563565/565992) and BD Horizon™ BV421 Rat Anti-Mouse CD25 (Cat. No. 564370) antibodies and with either BD Horizon™ RB780 Rat IgG2a, κ Isotype Control (Middle Plot) or BD Horizon™ RB780 Rat Anti-Mouse CD326 antibody (Right Plot) at 1.0 μg/test. The bivariate pseudocolor density plot showing the correlated expression of CD326 (or Ig Isotype control staining) versus CD3e was derived from CD25+ gated events with the light-scatter characteristics of viable splenic leucocytes. A small population of CD3+CD25+CD326+ cells was detected (Right Plot), whereas the CD25- T cells did not express detectable levels of CD326 (data not shown). Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 SE Cell Analyzer System and FlowJo™ Software. Data shown on this Technical Data Sheet are not lot specific.
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Multiparameter flow cytometric analysis of CD326 expression on BALB/c Mouse thymocytes and splenic T lymphocytes. Left Plot: Mouse thymocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) [Cat. No. 553141/553142] and stained with BD OptiBuild™ BV480 Rat Anti-Mouse CD4 (Cat. No. 746475) and BD Horizon™ BUV737 Rat Anti-Mouse CD8a (Cat. No. 612759) antibodies and with either BD Horizon™ RB780 Rat IgG2a, κ Isotype Control (Cat. No. 568698; dashed line histogram) or BD Horizon™ RB780 Rat Anti-Mouse CD326 antibody (Cat. No. 568736/568737; solid line histogram) at 1.0 μg/test. The histogram showing CD326 expression (or Ig Isotype control staining) was derived from CD4- and CD8-negative gated events with the forward and side light-scatter characteristics of viable thymocytes. Middle and Right Plots: Mouse splenic leucocytes were preincubated with Mouse BD Fc Block™ and stained with BD Horizon™ BUV395 Hamster Anti-Mouse CD3e (Cat. No. 563565/565992) and BD Horizon™ BV421 Rat Anti-Mouse CD25 (Cat. No. 564370) antibodies and with either BD Horizon™ RB780 Rat IgG2a, κ Isotype Control (Middle Plot) or BD Horizon™ RB780 Rat Anti-Mouse CD326 antibody (Right Plot) at 1.0 μg/test. The bivariate pseudocolor density plot showing the correlated expression of CD326 (or Ig Isotype control staining) versus CD3e was derived from CD25+ gated events with the light-scatter characteristics of viable splenic leucocytes. A small population of CD3+CD25+CD326+ cells was detected (Right Plot), whereas the CD25- T cells did not express detectable levels of CD326 (data not shown). Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 SE Cell Analyzer System and FlowJo™ Software. Data shown on this Technical Data Sheet are not lot specific.
Product Details
BD Horizon™
Ep-CAM; EGP; EGP-2; Egp314; GA733-2; TROP1; Tacsd1; Tacstd1; Ly74; gp40
Mouse (QC Testing)
Rat SD, also known as Sprague-Dawley (outbred) IgG2a, κ
Glycoconjugates from BALB/c mouse-derived TE-71 medullary thymic epithelial cell line
Flow cytometry (Routinely Tested)
0.2 mg/ml
17075
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO
Preparation And Storage
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to the dye under optimum conditions and unreacted dye was removed.
Recommended Assay Procedures
BD® CompBeads can be used as surrogates to assess fluorescence spillover (compensation). When fluorochrome conjugated antibodies are bound to BD® CompBeads, they have spectral properties very similar to cells. However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls. It is strongly recommended that when using a reagent for the first time, users compare the spillover on cells and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.
Product Notices
- When using high concentrations of antibody, background binding of this dye to erythroid fragments produced by ammonium chloride-based lysis, such as with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899), has been observed when the antibody conjugate was present during the lysis procedure. This may cause nonspecific staining of target cells, such as leukocytes, which have bound the resulting erythroid fragments. This background can be mitigated by any of the following: titrating the antibody conjugate to a lower concentration, fixing samples with formaldehyde, or removing erythrocytes before staining (eg, gradient centrifugation or pre-lysis with wash). This background has not been observed when cells were lysed with BD FACS™ Lysing Solution (Cat. No. 349202) after staining.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- An isotype control should be used at the same concentration as the antibody of interest.
- Please observe the following precautions: Absorption of visible light can significantly alter the energy transfer occurring in any tandem fluorochrome conjugate; therefore, we recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to prevent exposure of conjugated reagents, including cells stained with those reagents, to room illumination.
- Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- Cy is a trademark of Global Life Sciences Solutions Germany GmbH or an affiliate doing business as Cytiva.
Companion Products
Stain Buffer (FBS) RUO
Size
500 mL
Cat No.
554656
Purified Rat Anti-Mouse CD16/CD32 (Mouse BD Fc Block™) RUO
Size
0.5 mg
Cat No.
553142
RB780 Rat IgG2a, κ Isotype Control RUO
Size
50 µg
Cat No.
568698
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BV480 Rat Anti-Mouse CD4 RUO
Size
50 µg
Cat No.
746475
BUV737 Rat Anti-Mouse CD8a RUO
Size
50 µg
Cat No.
612759
BUV395 Hamster Anti-Mouse CD3e RUO
Size
50 µg
Cat No.
563565
568737 Rev. 2
Antibody Details
G8.8
The G8.8 monoclonal antibody recognizes CD326/Ep-CAM (Epithelial Cell Adhesion Molecule), also known as gp40 in the mouse and by a variety of names (including GA733-2, CO17-1A, and EGP) in the human. In the mouse, Ep-CAM is a 40-42 kDa cell-surface type 1 transmembrane glycoprotein expressed on thymic epithelial cells, thymic dendritic cells, immature thymocytes, a small subset of peripheral T lymphocytes, intestinal epithelium, kidney-collecting tubule epithelium, keratinocytes, Langerhans cells and lymph node and splenic dendritic cells. Profiles of Ep-CAM expression on fetal thymocytes and on the CD4[-] CD8[-], CD4[+] CD8[+], CD4[-] CD8[+], and CD4[+] CD8[-] subsets of adult thymocytes have been published. In unrelated studies, mouse Ep-CAM mRNA was detected in tissues containing epithelial cells (kidney, stomach, intestine, lung, and thymus) and in plasma cells and plasmacytomas, but not in heart, muscle, liver, brain, spleen, B lymphomas, or pre-B lymphomas. Ep-CAM is a Ca[2+] independent homophilic adhesion molecule that is proposed to play roles in the development and normal function of epithelial tissues and in the progression of carcinomas.
568737 Rev. 2
Format Details
RB780
The BD Horizon RealBlue™ 780 (RB780) Dye is part of the BD family of blue dyes. It is a tandem fluorochrome with an excitation maximum (Ex Max) at 498-nm and an emission maximum (Em Max) at 781-nm. Driven by BD innovation, RB780 can be used on both spectral and conventional cytometers and is designed to be excited by the Blue laser (488-nm) with minimal excitation by the 561-nm Yellow-Green laser. For conventional instruments equipped with a Blue laser (488-nm), RB780 can be used as an alternative to PE-Cy7 and we recommend using an optical filter centered near 780-nm (eg, a 780/60-nm bandpass filter). For spectral instruments equipped with a Blue laser (488-nm), it can be used in conjunction with PE-Cy7. RB780 is on average brighter than PE-Cy7 and has minimal spillover into Yellow-Green detectors.
RB780
Blue 488 nm
498 nm
781 nm
568737 Rev.2
Citations & References
Development References (9)
-
Basak S, Speicher D, Eck S, Wunner W. Colorectal carcinoma invasion inhibition by CO17-1A/GA733 antigen and its murine homologue. J Natl Cancer Inst. 1998; 90(9):691-697. (Biology). View Reference
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Bergsagel PL, Victor-Kobrin C, Timblin CR, Trepel J, Kuehl WM. A murine cDNA encodes a pan-epithelial glycoprotein that is also expressed on plasma cells. J Immunol. 1992; 148(2):590-596. (Biology). View Reference
-
Birebent B, Somasundaram R, Purev E. Anti-idiotypic antibody and recombinant antigen vaccines in colorectal cancer patients. Crit Rev Oncol Hematol. 2001; 39((1-2)):107-113. (Biology). View Reference
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Borkowski TA, Nelson AJ, Farr AG, Udey MC. Expression of gp40, the murine homologue of human epithelial cell adhesion molecule (Ep-CAM), by murine dendritic cells. Eur J Immunol. 1996; 26(1):110-114. (Clone-specific: Immunohistochemistry). View Reference
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Farr A, Nelson A, Truex J, Hosier S. Epithelial heterogeneity in the murine thymus: a cell surface glycoprotein expressed by subcapsular and medullary epithelium. J Histochem Cytochem. 1991; 39(5):345-353. (Immunogen: Electron microscopy, Immunohistochemistry, Immunoprecipitation). View Reference
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Litvinov SV, Balzar M, Winter MJ. Epithelial cell adhesion molecule (Ep-CAM) modulates cell-cell interactions mediated by classic cadherins. J Biol Chem. 1997; 139(5):1337-1348. (Biology). View Reference
-
Nelson AJ, Dunn RJ, Peach R, Aruffo A, Farr AG. The murine homolog of human Ep-CAM, a homotypic adhesion molecule, is expressed by thymocytes and thymic epithelial cells. Eur J Immunol. 1996; 26(2):401-408. (Clone-specific: Electron microscopy, Immunohistochemistry, Immunoprecipitation). View Reference
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Taguchi N, Hashimoto Y, Naiki M. Abnormal thymic expression of epithelial cell adhesion molecule (EP-CAM) in New Zealand Black (NZB) mice. J Autoimmun. 1999; 13(4):393-400. (Clone-specific: Electron microscopy). View Reference
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Zutter MM. Gastrointestinal carcinoma antigen GA733: target for immunodestruction and potential modifier of invasiveness and chemoresponsiveness. J Natl Cancer Inst. 1998; 90(9):642-644. (Biology). View Reference
568737 Rev. 2
Please refer to Support Documents for Quality Certificates
Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described
Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims. Comparisons are not made against non-BD technologies, unless otherwise noted.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.
